J Nutr Health.  2016 Apr;49(2):72-79. 10.4163/jnh.2016.49.2.72.

Elevated folic acid results in contrasting cancer cell line growth with implications for mandatory folic acid fortification

Affiliations
  • 1School of Biomedical Sciences and Pharmacy, University of Newcastle, Ourimbah NSW 2258, Australia.
  • 2School of Environmental & Life Sciences, University of Newcastle, Ourimbah NSW 2258, Australia. 73805@ncc.re.kr
  • 3Teaching & Research Unit, Central Coast Local Health District, Gosford NSW 2250, Australia.

Abstract

PURPOSE
The initiation of mandatory folic acid fortification using pteroylmonoglutamic acid (PteGlu) has reduced the rate of congenital malformations. However, it also appears to be responsible for several adverse effects, including increased cancer incidence. This may be related to physicho-chemical characteristics of PteGlu. This study examines the potential effect of high concentrations of PteGlu on a population subjected to mandatory folic acid fortification using an in vitro model.
METHODS
Caco-2 (colorectal cancer) and MCF7 (breast cancer) cell lines were cultured at 6 different PteGlu concentrations (0, 0.1, 1, 50, 250, and 500µg/ml) for 6 days. Cell growth was determined using thiazolyl blue tetrazolium bromide assay. The genotype of dihydrofolate reductase 19bp deletion/insertion (DHFR 19-del) was also scored in cell lines using a restriction fragment length polymorphism technique to examine whether genetic variations may factor in cell proliferation.
RESULTS
PteGlu exhibited differential growth promoting properties between cell lines. Caco-2 cells did not show a significant growth difference at low concentrations compared to control, however, at higher concentrations, the growth showed a contrasting trend in the early experimental period, while MCF7 showed enhanced cell growth at all concentrations. The DHFR 19-del genotype differed in the two cell lines.
CONCLUSION
Altered response to PteGlu by Caco-2 and MCF7 may reflect a tissue specific disease aetiology or genotype specific differential enzyme activity, for example by DHFR, to critical levels of PteGlu. As folic acid fortification is a blanket intervention, and DHFR and other enzyme activities vary between individuals, PteGlu intake may have an as yet undefined effect on health. These findings may be relevant when considering mandatory folic acid fortification for disease prevention.

Keyword

folate; Caco-2; MCF7; DHFR; folic acid fortification

MeSH Terms

Caco-2 Cells
Cell Line*
Cell Proliferation
Folic Acid*
Genetic Variation
Genotype
Humans
Incidence
Polymorphism, Restriction Fragment Length
Tetrahydrofolate Dehydrogenase
Folic Acid
Tetrahydrofolate Dehydrogenase

Figure

  • Fig. 1 Effects of PteGlu on growth of colon cancer cell line (Caco-2) for 6 days. Caco-2 cells were seeded at a density of 3×103/well and were cultured in medium with the presence of 5 different PteGlu concentrations, including a control (0 µg/ml). Cell growth was determined on day 2, 4 and 6, using thiazolyl blue tetrazolium bromide assay, and was presented as a percent of control. Each bar expresses the mean ± SD from triplicate analyses. Asterisks present the significant differences compared to the control group as determined by ANOVA with a confidence level of 95% (***p = 0.008 and ***p = 0.005). 1) PteGlu: pteroylmonoglutamic acid

  • Fig. 2 Effects of PteGlu on growth of breast cancer cell line (MCF7) for 6 days. MCF7 cells were seeded at a density of 3×103/well and were cultured in medium with the presence of 5 different PteGlu concentrations, including a control (0 µg/ml). Cell growth was determined on day 2, 4 and 6, using thiazolyl blue tetrazolium bromide assay, and was presented as a percent of control. Each bar expresses the mean ± SD from triplicate analyses. Asterisks present the significant differences compared to the control group as determined by ANOVA with a confidence level of 95% (***p < 0.0001). 1) PteGlu: pteroylmonoglutamic acid

  • Fig. 3 Scheme for folate metabolism showing utilisation of PteGlu and flux of one carbon units between DNA synthesis and methylation. Abbreviations: DHFR, dihydrofolate reductase; H2PteGlu, dihydrofolate; H4PteGlu, tetrahydrofolate; SHMT, serine hydroxymethyltransferase; MTHFR, methylentetrahydrofolate reductase; MS, methionine synthase; MSR, methyionine synthase reductase; BHMT, betainehomocysteine methyltransferase; Hcy, homocysteine; DMG, dymethylglycine; SAM, S-adenosylmethionine; SAH, S-adenosylhomocysteine

  • Fig. 4 Reaction diagram for PteGlu and dihydrofolate reductase. Abbreviations: DHFR, dihydrofolate reductase; H2PteGlu, dihydrofolate; H4PteGlu, tetrahydrofolate


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